Part of the ongoing research of the laboratory is to further understand the role of alpha6beta4 integrin in cellular physiologic processes as well as in growth factor-related transduction events. To explore the role of alpha6beta4 in these processes, we have recently developed mice carrying a targeted deletion of the C-terminal segment of the cytoplasmic tail of beta4 integrin (beta4 knock-in mice). This beta4 segment contains all phosphorylation sites that recruit the adapter protein Shc. Preliminary results suggest that alpha6beta4 integrin is involved in postnatal and tumor angiogenesis. The experiments described in this proposal have been designed to evaluate the importance of alpha6beta4 integrin in these processes and delineate the signaling pathways that alpha6beta4 participates. Angiogenesis will be evaluated by several methods such as by injection of matrigel plugs supplemented with growth factors, by the aortic ring assay, by subcutaneous injection of cancer cells or by exposure of young mice in hypoxic conditions and examining their retinas. In addition, beta4 integrin signaling will be examined in the knock-in mice using primary endothelial cells previously isolated from the lungs of these animals. Cellular physiologic processes which are important during postnatal and pathological angiogenesis such as cell migration, adhesion, proliferation and survival will be examined in primary endothelial cells of the beta4 knock-in mouse relative to wild-type. Transfection of these cells with dominant negative forms of signaling molecules downstream of beta4 integrin will be employed to determine the specific signaling pathways that become affected from impairment of alpha6beta4 signaling. The role of alpha6beta4 in tumor angiogenesis will also be evaluated in human glioblastoma cells that would be implanted in beta4 knock-in mice or induced by retroviral infection of glial precursor cells in the beta4 knock-in mice. The proposed research would be beneficial towards the understanding of both alpha6beta4 integrin function as well as the molecular basis of postnatal and pathological angiogenesis.